Project Summary/Abstract Seven million people suffer bone fractures annually in the U.S. Musculoskeletal conditions cost $215 billion/year. These numbers are increasing as the population ages. Calcium phosphate cement (CPC) can be molded and set in-situ to form hydroxyapatite, is osteoconductive, and can be resorbed and replaced by new bone. However, the low strength of CPC limits its use to non-stress locations. In the original five years of this grant, a new group of strong and macroporous CPCs were developed. Scaffolds with long macropores for tissue ingrowth and tailored strength history were generated. Currently, pre-fabricated carriers for stem cell delivery have difficulty in seeding cells deep into the scaffold, and cannot be injected in minimally-invasive procedures. Current injectable carriers are weak and cannot be used in a wide range of load-bearing repairs. Therefore, our objective for the next five years is to develop injectable, strong, tough, and macroporous nano- apatite scaffolds with stem cell and growth factor delivery for dental, craniofacial and orthopedic applications. In Aim 1, a new class of injectable, strong, tough and macroporous CPCs will be developed. The hypotheses are: (i) CPC composition can be tailored to improve injectability and strength; (ii) Optimizing the reinforcement and macroporosity will yield CPC with high-strain to accommodate for micro-motions within the tissues; (iii) The biomimetic nano-apatite scaffolds will enhance the colonization and differentiation of mesenchymal stem cells (MSCs) derived from rat bone marrow. Aim 2 will investigate growth factor delivery and test these hypotheses: (i) Fast-setting, strong and macroporous CPC-growth factor carrier can be formulated; (ii) Growth factor release from CPC is proportional to pore volume fraction; (iii) Controlled sequential release of multiple growth factors can be achieved to optimize stem cell function. Aim 3 will deliver stem cells and test these hypotheses: (i) Stem cells can be encapsulated in hydrogel and incorporated into CPC without decreasing cell viability and differentiation; (ii) Hydrogel beads can dissolve to release the cells and concomitantly create interconnected macropores in CPC; (iii) Stem cells and growth factors can be co-delivered in the same carrier to enhance cell function. Aim 4 will evaluate bone regeneration in animal models and test these hypotheses: (i) Macroporous CPC delivering stem cells and osteogenic and angiogenic growth factors will be completely resorbed and replaced by new bone across the entire critical-sized cranial defect in rats; (ii) The injectable, strong and macroporous CPCs have much higher resorption and new bone formation rates than traditional CPC; (iii) Optimizing the scaffold composition, macroporosity, and multiple growth factors will greatly enhance bone formation via stem cells. This new generation of injectable, strong and macroporous nano-apatite scaffolds with stem cell and growth factor delivery are expected to have dental, craniofacial and orthopedic applications, with greatly enhanced bone regeneration to improve the health and quality of life for millions of people.